Frontiers in Aging Neuroscience.  2013 May; 5(19):1-9

Thrombin, a mediator of cerebrovascular inflammation in AD and hypoxia.

Tripathy D, Sanchez A, Yin X, Luo J, Martinez J, Grammas P.

Garrison Institute on Aging, Texas Tech University Health Sciences Center, Lubbock, Texas 79430, USA.



Considerable evidence implicates hypoxia and vascular inflammation in Alzheimer’s disease (AD). Thrombin, a multifunctional inflammatory mediator, is demonstrable in the brains of AD patients both in the vessel walls and senile plaques. Hypoxia-inducible factor 1α (HIF-1α), a key regulator of the cellular response to hypoxia, is also upregulated in the vasculature of human AD brains. The objective of this study is to investigate inflammatory protein expression in the cerebrovasculature of transgenic AD mice and to explore the role of thrombin as a mediator of cerebrovascular inflammation and oxidative stress in AD and in hypoxia-induced changes in brain endothelial cells. Immunofluorescent analysis of the cerebrovasculature in AD mice demonstrates significant (p < 0.01-0.001) increases in thrombin, HIF-1α, interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), matrix metalloproteinases (MMPs), and reactive oxygen species (ROS) compared to controls. Administration of the thrombin inhibitor dabigatran (100 mg/kg) to AD mice for 34 weeks significantly decreases expression of inflammatory proteins and ROS. Exposure of cultured brain endothelial cells to hypoxia for 6 h causes an upregulation of thrombin, HIF-1α, MCP-1, IL-6, and MMP2 and ROS. Treatment of endothelial cells with the dabigatran (1 nM) reduces ROS generation and inflammatory protein expression (p < 0.01-0.001). The data demonstrate that inhibition of thrombin in culture blocks the increase in inflammatory protein expression and ROS generation evoked by hypoxia. Also, administration of dabigatran to transgenic AD mice diminishes ROS levels in brain and reduces cerebrovascular expression of inflammatory proteins. Taken together, these results suggest that inhibiting thrombin generation could have therapeutic value in AD and other disorders where hypoxia, inflammation, and oxidative stress are involved.

PMID: 23675346



Background Information:  Alzheimer’s disease (AD) is a progressive, irreversible, neurodegenerative disease age related disease.   The global societal burden of Alzheimer’s disease (AD) is astonishing. As of 2010, 36 million people world-wide have AD with 115 million expected to develop the disease by 2050.  Current healthcare expenditures exceed $604 billion [1]. At present, the few agents that are FDA-approved for the treatment of AD have demonstrated only modest effects in modifying clinical symptoms for relatively short periods and none have shown a clear effect on disease progression.  New therapeutic approaches are desperately needed.  In this project we postulate a new target for therapeutic intervention in AD, the activated/altered cerebral vascular endothelium.  There is a growing literature suggesting that perturbations in cerebral vascular structure and function occur in AD [2-8].  We hypothesize that in response to a persistent stimulus, brain endothelial cells become activated. Despite the continued presence of the stimulus, an imbalance of pro- and anti-angiogenic factors or aborted angiogenic signaling prevents new vessel growth. Therefore, in the absence of feedback signals to shut off vascular activation, endothelial cells continue to produce a large number of proteases, inflammatory proteins and other gene products with biologic activity that could injure or kill neurons.  Work from our laboratory has demonstrated a dysfunctional cerebral microcirculation in AD characterized by the upregulation of numerous inflammatory proteins and neurotoxic factors including vascular endothelial growth factor (VEGF) and thrombin [9-15].  Many of these vascular-derived factors are directly injurious to or lethal for neurons.  Vascular-derived factors can also injure neurons indirectly by activating neighboring glial cells which in turn release reactive oxygen species and inflammatory factors thus propagating deleterious neuroinflammation [16].

At first glance inhibiting vascular activation in AD may seem counterintuitive. We have shown that human brain microvessels derived from AD patients express or release inflammatory proteins and proteases, all of which have been implicated in endothelial activation and angiogenesis. Despite increases in these pro-activation/angiogenic factors in the AD brain, evidence for increased vascularity is lacking. On the contrary, it has been suggested that the angiogenic process is delayed and/or impaired in aged tissues [17-20] and that amyloid beta peptides have anti-angiogenic effects in vitro and in vivo [20]. How can the data showing anti-angiogenic effects of amyloid beta be reconciled with the presence or expression of a large number of proactivation/angiogenic proteins by brain microvessels in AD? In our working model, we hypothesize that in response to a persistent stimulus, such as cerebral hypoperfusion, one of the major clinical features in AD, brain endothelial cells become activated, acquiring an “activated/altered phenotype (Figure 1).”

Despite the continued presence of the stimulus, the multi-step process of angiogenesis leading to new blood vessel formation does not occur. Why vascular activation does not progress is unknown.  Therefore, in the absence of the end product, i.e. vessel growth, there are no feedback signals to shut off vascular activation. Endothelial cells remain activated and elaborate a large number of proteases, inflammatory proteins and other gene products with biologic activity that could injure or kill neurons. The “activated” brain vasculature represents an important and unexplored source of neurotoxins in the AD brain.

Thrombin, a multi-functional serine protease, is a key player in vascular activation.  Thrombin appears to work synergistically with VEGF and other factors released in AD [21] causing more damage in concert with other angiogenic and inflammatory factors than by itself, and idea furthered by the fact that neuronal cell death caused by thrombin or matrix metalloproteinase-9 is greater when both are present [22].  Targeting multi-functional factors such as thrombin could offer new therapeutic avenues for the treatment of so far intractable neurodegenerative diseases such as AD.

Joseph M. Martinez-1

Figure 1.  A proposed scheme for vascular activation as a contributing mechanism to neuronal injury in Alzheimer’s disease. Stress, such as hypoxia, serves as a trigger for endothelial cell activation in angiogenesis.  In the absence of new blood vessels to terminate the angiogenic process, the activated vasculature continually releases inflammatory proteins, neurotoxins and proteases that are directly neurotoxic and/or can potentiate inflammation of endothelial cells and the release of reactive oxygen species (ROS) by neighboring cells such as microglia and astrocytes.  Targeting thrombin using the direct thrombin inhibitor (DTI) dabigatran offers a novel way to mitigate the neuronal injury seen in Alzheimer’s disease by disrupting the release of thrombin and subsequent signal amplification.



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